Inbred maize line 413A

ABSTRACT

An inbred maize line, designated 413A, having higher row number and smaller kernel width compared to Ia2132, and herbicide tolerance, the plants and seeds of inbred maize line 413A and descendants thereof, methods for producing a maize plant produced by crossing the inbred line 413A with itself or with another maize plant, and hybrid maize seeds and plants produced by crossing the inbred line 413A with another maize line or plant.

FIELD OF THE INVENTION

This invention is in the field of maize breeding, specifically relatingto an inbred maize line designated 413A.

BACKGROUND OF THE INVENTION

The goal of plant breeding is to combine in a single variety or hybridvarious desirable traits. For field crops, these traits may includeresistance to diseases and insects, tolerance to heat and drought,reducing the time to crop maturity, greater yield, and better agronomicquality. With mechanical harvesting of many crops, uniformity of plantcharacteristics such as germination and stand establishment, growthrate, maturity, and plant and ear height, is important.

Field crops are bred through techniques that take advantage of theplant's method of pollination. A plant is self-pollinated if pollen fromone flower is transferred to the same or another flower of the sameplant. A plant is cross-pollinated if the pollen comes from a flower ona different plant. Plants that have been self-pollinated and selectedfor type for many generations become homozygous at almost all gene lociand produce a uniform population of true breeding progeny. A crossbetween two different homozygous lines produces a uniform population ofhybrid plants that may be heterozygous for many gene loci. A cross oftwo plants each heterozygous at a number of gene loci will produce apopulation of hybrid plants that differ genetically and will not beuniform.

Maize (Zea mays L.), often referred to as corn in the United States, canbe bred by both self-pollination and cross-pollination techniques. Maizehas separate male and female flowers on the same plant, located on thetassel and the ear, respectively. Natural pollination occurs in maizewhen wind blows pollen from the tassels to the silks that protrude fromthe tops of the ears.

A reliable method of controlling male fertility in plants offers theopportunity for improved plant breeding. This is especially true fordevelopment of maize hybrids, which relies upon some sort of malesterility system. There are several options for controlling malefertility available to breeders, such as: manual or mechanicalemasculation (or detasseling), cytoplasmic male sterility, genetic malesterility, gametocides and the like.

Hybrid maize seed is typically produced by a male sterility systemincorporating manual or mechanical detasseling. Alternate strips of twomaize inbreds are planted in a field, and the pollen-bearing tassels areremoved from one of the inbreds (female). Providing that there issufficient isolation from sources of foreign maize pollen, the ears ofthe detasseled inbred will be fertilized only from the other inbred(male) and the resulting seed is therefore hybrid and will form hybridplants.

The laborious, and occasionally unreliable, detasseling process can beavoided by using cytoplasmic male-sterile (CMS) inbreds. Plants of a CMSinbred are male sterile as a result of factors resulting from thecytoplasmic, as opposed to the nuclear, genome. Thus, thischaracteristic is inherited exclusively through the female parent inmaize plants, since only the female provides cytoplasm to the fertilizedseed. CMS plants are fertilized with pollen from another inbred that isnot male-sterile. Pollen from the second inbred may or may notcontribute genes that make the hybrid plants male-fertile. Seed fromdetasseled fertile maize and CMS produced seed of the same hybrid can beblended to insure that adequate pollen loads are available forfertilization when the hybrid plants are grown.

There are several methods of conferring genetic male sterilityavailable, such as multiple mutant genes at separate locations withinthe genome that confer male sterility, as disclosed in U.S. Pat. Nos.4,654,465 and 4,727,219 and chromosomal translocations as described inU.S. Pat. Nos. 3,861,709 and 3,710,511, the disclosures of which arespecifically incorporated herein by reference. There are many othermethods of conferring genetic male sterility in the art, each with itsown benefits and drawbacks. These methods use a variety of approachessuch as delivering into the plant a gene encoding a cytotoxic substanceassociated with a male tissue specific promoter or an antisense systemin which a gene critical to fertility is identified and an antisense tothat gene is inserted in the plant (EPO 89/3010153.8 and WO 90/08828).

Another system useful in controlling male sterility makes use ofgametocides. Gametocides are not a genetic system, but rather a topicalapplication of chemicals. These chemicals affect cells that are criticalto male fertility. The application of these chemicals affects fertilityin the plants only for the growing season in which the gametocide isapplied (see Carlson, Glenn R., U.S. Pat. No. 4,936,904, which isincorporated herein by reference). Application of the gametocide, timingof the application and genotype specificity often limit the usefulnessof the approach.

The use of male sterile inbreds is but one factor in the production ofmaize hybrids. The development of maize hybrids requires, in general,the development of homozygous inbred lines, the crossing of these lines,and the evaluation of the crosses. Pedigree breeding and recurrentselection breeding methods are used to develop inbred lines frombreeding populations. Breeding programs combine the genetic backgroundsfrom two or more inbred lines or various other germplasm sources intobreeding pools from which new inbred lines are developed by selfing andselection of desired phenotypes. The new inbreds are crossed with otherinbred lines and the hybrids from these crosses are evaluated todetermine which of those have commercial potential. Plant breeding andhybrid development are expensive and time-consuming processes.

Pedigree breeding starts with the crossing of two genotypes, each ofwhich may have one or more desirable characteristics that is lacking inthe other or which complements the other. If the two original parents donot provide all the desired characteristics, other sources can beincluded in the breeding population. In the pedigree method, superiorplants are selfed and selected in successive generations. In thesucceeding generations the heterozygous condition gives way tohomogeneous lines as a result of self-pollination and selection.Typically in the pedigree method of breeding five or more generations ofselfing and selection is practiced: F1 to F2; F3 to F4; F4 to F5, etc.

Recurrent selection breeding can be used to improve populations ofeither self or cross-pollinating crops. Recurrent selection can be usedto transfer a specific desirable trait from one inbred or source to aninbred that lacks the trait. This can be accomplished, for example, byfirst a superior inbred (recurrent parent) to a donor inbred(non-recurrent parent), that carries the appropriate gene(s) for thetrait in question. The progeny of this cross is then mated back to thesuperior recurrent parent followed by selection in the resultant progenyfor the desired trait to be transferred from the non-recurrent parent.After five or more backcross generations with selection for the desiredtrait, the progeny will be homozygous for loci controlling thecharacteristic being transferred, but will be like the superior parentfor essentially all other genes. The last backcross generation is, thenselfed to give pure breeding progeny for the gene(s) being transferred.A hybrid developed from inbreds containing the transferred gene(s) isessentially the same as a hybrid developed form the same inbreds withoutthe transferred genes. As the varieties developed using recurrentselection breeding contain almost all of the characteristics of therecurrent parent, selecting a superior recurrent parent is desirable.

A single cross maize hybrid results from the cross of two inbred lines,each of which has a genotype that complements the genotype of the other.The hybrid progeny of the first generation is designated F1. In thedevelopment of commercial hybrids only the F1 hybrid plants are sought.Preferred F1 hybrids are more vigorous than their inbred parents. Thishybrid vigor, or heterosis, can be manifested in many polygenic traits,including increased vegetative growth and increased yield.

The development of a maize hybrid involves three steps: (1) theselection of plants from various germplasm pools for initial breedingcrosses; (2) the selfing of the selected plants from the breedingcrosses for several generations to produce a series of inbred lines,which, although different from each other, breed true and are highlyuniform; and (3) crossing the selected inbred lines with differentinbred lines to produce the hybrid progeny (F1). During the inbreedingprocess in maize, the vigor of the lines decreases. Vigor is restoredwhen two different inbred lines are crossed to produce the hybridprogeny (F1). An important consequence of the homozygosity andhomogeneity of the inbred lines is that the hybrid between a definedpair of inbreds will always be the same. Once the inbreds that give asuperior hybrid have been identified, the hybrid seed can be reproducedindefinitely as long as the homogeneity of the inbred parents ismaintained.

A single cross hybrid is produced when two inbred lines are crossed toproduce the F1 progeny. A double cross hybrid is produced from fourinbred lines crossed in pairs (A×B and C×D) and then the two F1 hybridsare crossed again (A×B)×(C×D). Much of the hybrid vigor exhibited by F1hybrids is lost in the next generation (F2). Consequently, seed fromhybrids is not used for planting stock.

Hybrid seed production requires elimination or inactivation of pollenproduced by the female parent. Incomplete removal or inactivation of thepollen provides the potential for self-pollination. This inadvertentlyself-pollinated seed may be unintentionally harvested and packaged withhybrid seed. Once the seed is planted, it is possible to identify andselect these self-pollinated plants. These self-pollinated plants willbe genetically equivalent to the female inbred line used to produce thehybrid. Typically these self-pollinated plants can be identified andselected due to their decreased vigor. Female selfs are identified bytheir less vigorous appearance for vegetative and/or reproductivecharacteristics, including shorter plant height, small ear size, ear andkernel shape, cob color, or other characteristics.

Identification of these self-pollinated lines can also be accomplishedthrough molecular marker analyses. See, “The Identification of FemaleSelfs in Hybrid Maize: A Comparison Using Electrophoresis andMorphology”, Smith, J. S. C. and Wych, R. D., Seed Science andTechnology 14, pp. 1-8 (1995), the disclosure of which is expresslyincorporated herein by reference. Through these technologies, thehomozygosity of the self-pollinated line can be verified by analyzingallelic composition at various loci along the genome. Those methodsallow for rapid identification of the invention disclosed herein. Seealso, “Identification of Atypical Plants in Hybrid Maize Seed byPostcontrol and Electrophoresis” Sarca, V. et al., Probleme de GeneticaTeoritca si Aplicata Vol. 20 (1) p. 29-42.

As is readily apparent to one skilled in the art, the foregoingdescribes only two of the various ways by which the inbred can beobtained by those looking to use the germplasm. Other means areavailable, and the above examples are illustrative only.

Maize is an important and valuable field crop. Thus, a continuing goalof plant breeders is to develop high-yielding maize hybrids that areagronomically sound based on stable inbred lines. The reasons for thisgoal are obvious: to maximize the amount of grain produced with theinputs used and minimize susceptibility of the crop to pests andenvironmental stresses. To accomplish this goal, the maize breeder mustselect and develop superior inbred parental lines for producing hybrids.This requires identification and selection of genetically uniqueindividuals that occur in a segregating population. The segregatingpopulation is the result of a combination of crossover events plus theindependent assortment of specific combinations of alleles at many geneloci that results in specific genotypes. The probability of selectingany one individual with a specific genotype from a breeding cross isinfinitesimal due to the large number of segregating genes and theunlimited recombinations of these genes, some of which may be closelylinked. However, the genetic variation among individual progeny of abreeding cross allows for the identification of rare and valuable newgenotypes. These new genotypes are neither predictable nor incrementalin value, but rather the result of manifested genetic variation combinedwith selection methods, environments and the actions of the breeder.Thus, even if the entire genotypes of the parents of the breeding crosswere characterized and a desired genotype known, only a few, if any,individuals having the desired genotype may be found in a largesegregating F2 population. Typically, however, neither the genotypes ofthe breeding cross parents nor the desired genotype to be selected isknown in any detail. In addition, it is not known how the desiredgenotype would react with the environment. This genotype by environmentinteraction is an important, yet unpredictable, factor in plantbreeding. A breeder of ordinary skill in the art cannot predict thegenotype, how that genotype will interact with various climaticconditions or the resulting phenotypes of the developing lines, exceptperhaps in a very broad and general fashion. A breeder of ordinary skillin the art would also be unable to recreate the same line twice from thevery same original parents, as the breeder is unable to direct how thegenomes combine or how they will interact with the environmentalconditions. This unpredictability results in the expenditure of largeamounts of research resources in the development of a superior new maizeinbred line.

SUMMARY OF THE INVENTION

According to the invention, there is provided a novel inbred maize line,designated 413A having higher row number and smaller kernel widthcompared to Ia2132, and herbicide tolerance. This invention thus relatesto the seeds of inbred maize line 413A, to the plants of inbred maizeline 413A, and to methods for producing a maize plant by crossing theinbred line 413A with itself or another maize line. This inventionfurther relates to hybrid maize seeds and plants produced by crossingthe inbred line 413A with another maize line.

The invention is also directed to inbred maize line 413A into which oneor more specific, single gene traits, for example transgenes, have beenintrogressed from another maize line. Preferably, the resulting line hasessentially all of the morphological and physiological characteristicsof inbred maize line of 413A, in addition to the one or more specific,single gene traits introgressed into the inbred, preferably theresulting line has all of the morphological and physiologicalcharacteristics of inbred maize line of 413A, in addition to the one ormore specific, single gene traits introgressed into the inbred. Theinvention also relates to seeds of an inbred maize line 413A into whichone or more specific, single gene traits have been introgressed and toplants of an inbred maize line 413A into which one or more specific,single gene traits have been introgressed. The invention further relatesto methods for producing a maize plant by crossing plants of an inbredmaize line 413A into which one or more specific, single gene traits havebeen introgressed with themselves or with another maize line. Theinvention also further relates to hybrid maize seeds and plants producedby crossing plants of an inbred maize line 413A into which one or morespecific, single gene traits have been introgressed with another maizeline. The invention is also directed to a method of producing inbredscomprising planting a collection of hybrid seed, growing plants from thecollection, identifying inbreds among the hybrid plants, selecting theinbred plants and controlling their pollination to preserve theirhomozygosity.

DETAILED DESCRIPTION OF THE INVENTION

Inbred maize lines are typically developed for use in the production ofhybrid maize lines. Inbred maize lines need to be highly homogeneous,homozygous and reproducible to be useful as parents of commercialhybrids. There are many analytical methods available to determine thehomozygotic and phenotypic stability of these inbred lines.

The oldest and most traditional method of analysis is the observation ofphenotypic traits. The data is usually collected in field experimentsover the life of the maize plants to be examined. Phenotypiccharacteristics most often observed are for traits associated with plantmorphology, ear and kernel morphology, insect and disease resistance,maturity, and yield.

In addition to phenotypic observations, the genotype of a plant can alsobe examined. There are many laboratory-based techniques available forthe analysis, comparison and characterization of plant genotype; amongthese are Isozyme Electrophoresis, Restriction Fragment LengthPolymorphisms (RFLPs), Randomly Amplified Polymorphic DNAs (RAPDs),Arbitrarily Primed Polymerase Chain Reaction (AP-PCR), DNA AmplificationFingerprinting (DAF), Sequence Characterized Amplified Regions (SCARs),Amplified Fragment Length Polymorphisms (AFLPs), and Simple SequenceRepeats (SSRs) which are also referred to as Microsatellites.

Some of the most widely used of these laboratory techniques are IsozymeElectrophoresis and RFLPs as discussed in Lee, M., “Inbred Lines ofMaize and Their Molecular Markers,” The Maize Handbook,(Springer-Verlag, New York, Inc. 1994, at 423-432). IsozymeElectrophoresis is a useful tool in determining genetic composition,although it has relatively low number of available markers and the lownumber of allelic variants among maize inbreds. RFLPs have the advantageof revealing an exceptionally high degree of allelic variation in maizeand the number of available markers is almost limitless. Maize RFLPlinkage maps have been rapidly constructed and widely implemented ingenetic studies. One such study is described in Boppenmaier, et al.,“Comparisons among strains of inbreds for RFLPs”, Maize GeneticsCooperative Newsletter, 65:1991, pg. 90. This study used 101 RFLPmarkers to analyze the patterns of 2 to 3 different deposits each offive different inbred lines. The inbred lines had been selfed from 9 to12 times before being adopted into 2 to 3 different breeding programs.It was results from these 2 to 3 different breeding programs thatsupplied the different deposits for analysis. These five lines weremaintained in the separate breeding programs by selfing or sibbing androgueing off-type plants for an additional one to eight generations.After the RFLP analysis was completed, it was determined the five linesshowed 0-2% residual heterozygosity. Although this was a relativelysmall study, it can be seen using RFLPs that the lines had been highlyhomozygous prior to the separate strain maintenance.

The production of hybrid maize lines typically comprises planting inpollinating proximity seeds of, for example, inbred maize line 413A andof a different inbred parent maize plant, cultivating the seeds ofinbred maize line 413A and of said different inbred parent maize plantinto plants that bear flowers, emasculating the male flowers of inbredmaize line 413A or the male flowers of said different inbred parentmaize plant to produce an emasculated maize plant, allowingcross-pollination to occur between inbred maize line 413A and saiddifferent inbred parent maize plant and harvesting seeds produced onsaid emasculated maize plant. The harvested seed are grown to producehybrid maize plants.

Inbred maize line 413A can be crossed to inbred maize lines of variousheterotic group (see e.g. Hallauer et al. (1988) in Corn and CornImprovement, Sprague et al, eds, chapter 8, pages 463-564) for theproduction of hybrid maize lines. TABLE I VARIETY DESCRIPTIONINFORMATION Inbred maize line 413A is compared to inbred Ia2132 413AIa2132 Std LSD Mean Dev Mean Std Dev .05 Sig Y/N PLANT Plant height (cm)200.6 13.5 168.4 14.1 4.93 Y Ear height (cm) 76.2 9.8 40.0 17.0 3.63 YInternode length (cm) 14.0 2.0 14.6 2.2 0.78 N Number of tillers 1.7 1.02.0 0.6 0.32 Y Ears per stalk 1.1 0.2 1.5 0.8 0.24 Y LEAF Width of earnode leaf 5.4 0.5 5.9 1.3 0.41 Y (cm) Length of ear node leaf 82.8 0.560.9 8.7 2.99 Y (cm) Number of leaves above 4.6 0.5 5.3 0.7 0.23 Y Leafangle (degrees from 3.2 5.4 65.8 10.3 2.89 Y top of stalk) TASSEL Numberof Primary 11.9 3.0 18.1 2.6 1.19 Y Lateral Branches Branch Angle(degrees 75.4 11.3 35.8 4.4 3.67 Y from central spike) Tassel length(cm) 46.0 3.6 33.6 3.9 1.47 Y EAR Ear length (cm) 11.7 1.0 11.4 1.7 0.53N Ear diameter (cm) 38.1 3.0 36.3 3.3 1.28 Y Row number 23.2 2.1 12.71.1 0.74 Y Kernel length (mm) 11.6 0.9 10.7 1.4 0.43 Y Kernel width (mm)6.5 0.6 8.3 0.9 0.35 Y Kernel thickness (mm) 3.0 0.2 3.4 0.8 0.25 NPercentage of round 3.9 3.2 35.8 11.9 3.81 Y kernels Weight of 100kernels 11.1 2.1 19.5 2.4 1.02 Y (grams) Cob diameter (mm) 13.1 1.3 12.41.1 0.5 Y Descriptive Ratings 413A Ia2132 (According to the PVP form)Leaf sheath pubescence 4.0 2.0 Marginal waves 6.0 5.0 Longitudinalcreases 3.0 8.0 Pollen shed 3 3 Kernel rows 1 Row alignment 2 2 Eartaper 2 2 Aleurone color pattern 1 1 Endosperm type 1 (su) 1 (su)Anthocyanin of brace 1 1 roots MATURITY Days Heat Units Days Heat UnitsEmergence to 50% of 74 67 plants in silk Emergence to 50% of 67 64plants in pollen 50% silk to optimum 95 88 edible quality COLOR PVP CodeMunsell PVP Code Munsell Leaf 03 7.5gy4/4 03 7.5gy4/5 Anther 14 2.5r5/814 2.5r4/4 Glume 14 2.5r5/8 14 5r4/6 Silk 01 2.5gy8/6 14 5r5/6 Freshhusk 02 5gy6/6 02 5gy7/8

In interpreting the foregoing color designations, reference may be madeto the Munsell Glossy Book of Color, a standard color reference. Colorcodes: 1. light green, 2. medium green, 3. dark green, 4. very darkgreen, 5. green-yellow, 6. pale yellow, 7. yellow, 8. yellow-orange, 9.salmon, 10. pink-orange, 11. pink 12. light red, 13. cherry red, 14.red, 15. red and white, 16. pale purple, 17. purple, 18. colorless, 19.white, 20, white capped, 21. buff, 22. tan, 23. brown, 24. bronze, 25.variegated, 26. other.

413A differs from Ia2132 for several different traits. These traits are:

The Plant Height of 413A is 201 cm while the plant height of Ia2132 is168 cm.

The Length of Ear Node Leaf of 413A is 83 cm while the length of earnode leaf of Ia2132 is 61 cm.

The Leaf Angle of 413A is 3 degrees while the leaf angle of Ia2132 is 66degrees.

The Longitudinal Creases on 413A is rated a 3 and is significantlydifferent than Ia2132, which is rated a 8.

The 413A tassel has fewer branches than the Ia2132 tassel. 413A has 12Primary Tassel Branches and Ia2132 has 18. The Tassel Branch Angle of413A is 75 degrees while the tassel branch angle for Ia2132 is 36degrees.

The ear of 413A is also different than the Ia2132 ear. The Ear Height of413A is 76 cm while the ear height of Ia2132 is 40 cm. 413A ear has 4Percent Round Kernels as compared to 36% on Ia2132. TABLE II HybridGH-5704 has inbred 413A and 363B as parents. Hybrids GH-5703 is used forcomparison. GH-5704 is a Poast herbicide resistant version of GH-5703with the herbicide resistance coming from 413A. Mid Silk Ear Row numberHusk Tip Trial ID Location Year Date Length ave. (in) ave. length (cm)fill (cm) Hybrid: GH-5704 T02NMP Nampa ID 2002 64 8 19.7 1 0.5 T02MN1Stanton MN 2002 72 7.4 20.7 0.5 1 T02TCF Othello WA 2002 na 8.2 na 2 1T02SMP Pasco, WA 2002 na 8 na 3 0.5 T02WILL Salem, OR 2002 na 7.6 na 4 0T01NMP Nampa ID 2001 63 8.1 22 0.5 0 T01MN1 Stanton MN 2001 81 7.8 210.5 3 Average:   70.0 7.9 20.9 1.6 0.9 Hybrid: GH-5703 T02NMP Nampa ID2002 64 8.1 22 0.5 0.5 T02MN1 Stanton MN 2002 72 7.8 21 1 2 T02TCFOthello WA 2002 na 8 na 2 1 T02SMP Pasco, WA 2002 na 7.5 na 3 0.5T02WILL Salem, OR 2002 na 7.6 na 4 0 T01NMP Nampa ID 2001 64 8 19.3 1 1T01MN1 Stanton MN 2001 78 8 18.7 0 2.5 Average:   69.5 7.9 20.3 1.6 1.1

Mid silk date is the number of days from planting to 50% plants with earsilk. Husk length is centimeters of husk past ear tip. Tip fill iscentimeters of blank tip below tip of ear. Common rust is a scale of 0-9with 0 equals none and 9 equals most severe.

The invention also encompasses plants of inbred maize line 413A andparts thereof further comprising one or more specific, single genetraits which have been introgressed into inbred maize line 413A fromanother maize line. Preferably, one or more new traits are transferredto inbred maize line 413A, or, alternatively, one or more traits ofinbred maize line 413A are altered or substituted. The transfer (orintrogression) of the trait(s) into inbred maize line 413A is forexample achieved by recurrent selection breeding, for example bybackcrossing. In this case, inbred maize line 413A (the recurrentparent) is first crossed to a donor inbred (the non-recurrent parent)that carries the appropriate gene(s) for the trait(s) in question. Theprogeny of this cross is then mated back to the recurrent parentfollowed by selection in the resultant progeny for the desired trait(s)to be transferred from the non-recurrent parent. After three, preferablyfour, more preferably five or more generations of backcrosses with therecurrent parent with selection for the desired trait(s), the progenywill be heterozygous for loci controlling the trait(s) beingtransferred, but will be like the recurrent parent for most or almostall other genes (see, for example, Poehlman & Sleper (1995) BreedingField Crops, 4th Ed., 172-175; Fehr (1987) Principles of CultivarDevelopment, Vol. 1: Theory and Technique, 360-376).

The laboratory-based techniques described above, in particular RFLP andSSR, are routinely used in such backcrosses to identify the progenieshaving the highest degree of genetic identity with the recurrent parent.This permits to accelerate the production of inbred maize lines havingat least 90%, preferably at least 95%, more preferably at least 99%genetic identity with the recurrent parent, yet more preferablygenetically identical to the recurrent parent, and further comprisingthe trait(s) introgressed from the donor patent. Such determination ofgenetic identity is based on molecular markers used in thelaboratory-based techniques described above. Such molecular markers arefor example those known in the art and described in Boppenmaier, et al.,“Comparisons among strains of inbreds for RFLPs”, Maize GeneticsCooperative Newsletter (1991) 65, pg. 90, or those available from theUniversity of Missouri database and the Brookhaven laboratory database(see http://www.agron.missouri.edu). The last backcross generation isthen selfed to give pure breeding progeny for the gene(s) beingtransferred. The resulting plants have essentially all of themorphological and physiological characteristics of inbred maize line413A, in addition to the single gene trait(s) transferred to the inbred.The exact backcrossing protocol will depend on the trait being alteredto determine an appropriate testing protocol. Although backcrossingmethods are simplified when the trait being transferred is a dominantallele, a recessive allele may also be transferred. In this instance itmay be necessary to introduce a test of the progeny to determine if thedesired trait has been successfully transferred.

Many traits have been identified that are not regularly selected for inthe development of a new inbred but that can be improved by backcrossingtechniques or genetic transformation. Examples of traits transferred toinbred maize line 413A include, but are not limited to, waxy starch,herbicide tolerance, resistance for bacterial, fungal, or viral disease,insect resistance, enhanced nutritional quality, improved performance inan industrial process, altered reproductive capability, such as malesterility or male fertility, yield stability and yield enhancement.Other traits transferred to inbred maize line 413A are for theproduction of commercially valuable enzymes or metabolites in plants ofinbred maize line 413A.

Traits transferred to maize inbred line 413A are naturally occurringmaize traits, which are preferably introgressed into inbred maize line413A by breeding methods such as backcrossing, or are heterologoustransgenes, which are preferably first introduced into a maize line bygenetic transformation using genetic engineering and transformationtechniques well known in the art, and then introgressed into inbred line413A. Alternatively a heterologous trait is directly introduced intoinbred maize line 413A by genetic transformation. Heterologous, as usedherein, means of different natural origin or represents a non-naturalstate. For example, if a host cell is transformed with a nucleotidesequence derived from another organism, particularly from anotherspecies, that nucleotide sequence is heterologous with respect to thathost cell and also with respect to descendants of the host cell whichcarry that gene. Similarly, heterologous refers to a nucleotide sequencederived from and inserted into the same natural, original cell type, butwhich is present in a non-natural state, e.g. a different copy number,or under the control of different regulatory sequences. A transformingnucleotide sequence may comprise a heterologous coding sequence, orheterologous regulatory sequences. Alternatively, the transformingnucleotide sequence may be completely heterologous or may comprise anypossible combination of heterologous and endogenous nucleic acidsequences.

A transgene introgressed into maize inbred line 413A typically comprisesa nucleotide sequence whose expression is responsible or contributes tothe trait under the control of a promoter appropriate for the expressionof the nucleotide sequence at the desired time in the desired tissue orpart of the plant. Constitutive or inducible promoters are used. Thetransgene may also comprise other regulatory elements such as forexample translation enhancers or termination signals. In a preferredembodiment, the nucleotide sequence is the coding sequence of a gene andis transcribed and translated into a protein. In another preferredembodiment, the nucleotide sequence encodes an antisense RNA, a senseRNA that is not translated or only partially translated, a t-RNA, ar-RNA or a sn-RNA.

Where more than one trait are introgressed into inbred maize line 413A,it is preferred that the specific genes are all located at the samegenomic locus in the donor, non-recurrent parent, preferably, in thecase of transgenes, as part of a single DNA construct integrated intothe donor's genome. Alternatively, if the genes are located at differentgenomic loci in the donor, non-recurrent parent, backcrossing allows torecover all of the morphological and physiological characteristics ofinbred maize line 413A in addition to the multiple genes in theresulting maize inbred line.

The genes responsible for a specific, single gene trait are generallyinherited through the nucleus. Known exceptions are, e.g. the genes formale sterility, some of which are inherited cytoplasmically, but stillact as single gene traits. In a preferred embodiment, a heterologoustransgene to be transferred to maize inbred line 413A is integrated intothe nuclear genome of the donor, non-recurrent parent. In anotherpreferred embodiment, a heterologous transgene to be transferred to intomaize inbred line 413A is integrated into the plastid genome of thedonor, non-recurrent parent. In a preferred embodiment, a plastidtransgene comprises one gene transcribed from a single promoter or twoor more genes transcribed from a single promoter.

In a preferred embodiment, a transgene whose expression results orcontributes to a desired trait to be transferred to maize inbred line413A comprises a virus resistance trait such as, for example, a MDMVstrain B coat protein gene whose expression confers resistance to mixedinfections of maize dwarf mosaic virus and maize chlorotic mottle virusin transgenic maize plants (Murry et al. Biotechnology (1993)11:1559-64). In another preferred embodiment, a transgene comprises agene encoding an insecticidal protein, such as, for example, a crystalprotein of Bacillus thuringiensis or a vegetative insecticidal proteinfrom Bacillus cereus, such as VIP3 (see for example Estruch et al. NatBiotechnol (1997) 15:137-41). In a preferred embodiment, an insecticidalgene introduced into maize inbred line 413A is a CrylAb gene or aportion thereof, for example introgressed into maize inbred line 413Afrom a maize line comprising a Bt-11 event as described in U.S. Pat. No.6,114,608, which is incorporated herein by reference, or from a maizeline comprising a 176 event as described in Koziel et al. (1993)Biotechnology 11:194-200. In yet another preferred embodiment, atransgene introgressed into maize inbred line 413A comprises a herbicidetolerance gene. For example, expression of an altered acetohydroxyacidsynthase (AHAS) enzyme confers upon plants tolerance to variousimidazolinone or sulfonamide herbicides (U.S. Pat. No. 4,761,373). Inanother preferred embodiment, a non-transgenic trait conferringtolerance to imidazolinones is introgressed into maize inbred line 413A(e.g a “IT” or “IR” trait). U.S. Pat. No. 4,975,374, incorporated hereinby reference, relates to plant cells and plants containing a geneencoding a mutant glutamine synthetase (GS) resistant to inhibition byherbicides that are known to inhibit GS, e.g. phosphinothricin andmethionine sulfoximine. Also, expression of a Streptomyces bar geneencoding a phosphinothricin acetyl transferase in maize plants resultsin tolerance to the herbicide phosphinothricin or glufosinate (U.S. Pat.No. 5,489,520). U.S. Pat. No. 5,013,659, which is incorporated herein byreference, is directed to plants that express a mutant acetolactatesynthase (ALS) that renders the plants resistant to inhibition bysulfonylurea herbicides. U.S. Pat. No. 5,162,602 discloses plantstolerant to inhibition by cyclohexanedione and aryloxyphenoxypropanoicacid herbicides. The tolerance is conferred by an altered acetylcoenzyme A carboxylase(ACCase). U.S. Pat. No. 5,554,798 disclosestransgenic glyphosate tolerant maize plants, which tolerance isconferred by an altered 5-enolpyruvyl-3-phosphoshikimate (EPSP) synthasegene. U.S. Pat. No. 5,804,425 discloses transgenic glyphosate tolerantmaize plants, which tolerance is conferred by an EPSP synthase genederived from Agrobacterium tumefaciens CP-4 strain. Also, tolerance to aprotoporphyrinogen oxidase inhibitor is achieved by expression of atolerant protoporphyrinogen oxidase enzyme in plants (U.S. Pat. No.5,767,373). Another trait transferred to inbred maize line 413A conferstolerance to an inhibitor of the enzyme hydroxyphenylpyruvatedioxygenase (HPPD) and transgenes conferring such trait are, forexample, described in WO 9638567, WO 9802562, WO 9923886, WO 9925842, WO9749816, WO 9804685 and WO 9904021. All issued patents referred toherein are, in their entirety, expressly incorporated herein byreference.

In a preferred embodiment, a transgene transferred to maize inbred line413A comprises a gene conferring tolerance to a herbicide and at leastanother nucleotide sequence encoding another trait, such as for example,an insecticidal protein. Such combination of single gene traits is forexample a CrylAb gene and a bar gene.

Specific transgenic events introgressed into maize inbred line 413A canbe obtained through the list of Petitions of Nonregulated Status grantedby APHIS as of Oct. 12, 2000. For example, introgressed from glyphosatetolerant event GA21 (9709901p), glyphosate tolerant/Lepidopteran insectresistant event MON 802 (9631701p), Lepidopteran insect resistant eventDBT418 (9629101p), male sterile event MS3 (9522801p), Lepidopteraninsect resistant event Bt11 (9519501p), phosphinothricin tolerant eventB16 (9514501p), Lepidopteran insect resistant event MON 80100(9509301p), phosphinothricin tolerant events T14, T25 (9435701p),Lepidopteran insect resistant event 176 (9431901p).

The introgression of a Bt11 event into a maize line, such as maizeinbred line 413A, by backcrossing is exemplified in U.S. Pat. No.6,114,608, and the present invention is directed to methods ofintrogressing a Bt11 event into maize inbred line 413A using for examplethe markers described in U.S. Pat. No. 6,114,608 and to resulting maizelines.

Direct selection may be applied where the trait acts as a dominanttrait. An example of a dominant trait is herbicide tolerance. For thisselection process, the progeny of the initial cross are sprayed with theherbicide prior to the backcrossing. The spraying eliminates any plantwhich does not have the desired herbicide tolerance characteristic, andonly those plants that have the herbicide tolerance gene are used in thesubsequent backcross. This process is then repeated for the additionalbackcross generations.

This invention also is directed to methods for producing a maize plantby crossing a first parent maize plant with a second parent maize plantwherein either the first or second parent maize plant is a maize plantof inbred line 413A or a maize plant of inbred line 413A furthercomprising one or more single gene traits. Further, both first andsecond parent maize plants can come from the inbred maize line 413A oran inbred maize plant of 413A further comprising one or more single genetraits. Thus, any such methods using the inbred maize line 413A or aninbred maize plant of 413A further comprising one or more single genetraits are part of this invention: selfing, backcrosses, hybridproduction, crosses to populations, and the like. All plants producedusing inbred maize line 413A or inbred maize plants of 413A firthercomprising one or more single gene traits as a parent are within thescope of this invention. Advantageously, inbred maize line 413A orinbred maize plants of 413A further comprising one or more single genetraits are used in crosses with other, different, maize inbreds toproduce first generation (F1) maize hybrid seeds and plants withsuperior characteristics.

In a preferred embodiment, seeds of inbred maize line 413A or seeds ofinbred maize plants of 413A further comprising one or more single genetraits are provided as an essentially homogeneous population of inbredcorn seeds. Essentially homogeneous populations of inbred seed are thosethat consist essentially of the particular inbred seed, and aregenerally purified free from substantial numbers of other seed, so thatthe inbred seed forms between about 90% and about 100% of the totalseed, and preferably, between about 95% and about 100% of the totalseed. Most preferably, an essentially homogeneous population of inbredcorn seed will contain between about 98.5%, 99%, 99.5% and about 100% ofinbred seed, as measured by seed grow outs. The population of inbredcorn seeds of the invention is further particularly defined as beingessentially free from hybrid seed. The inbred seed population may beseparately grown to provide an essentially homogeneous population ofplants of inbred maize line 413A or inbred maize plants of 413A furthercomprising one or more single gene traits.

As used herein, the term “plant” includes plant cells, plantprotoplasts, plant cell tissue cultures from which maize plants can beregenerated, plant calli, plant clumps, and plant cells that are intactin plants or parts of plants, such as embryos, pollen, ovules, flowers,kernels, ears, cobs, leaves, husks, stalks, roots, root tips, anthers,silk, seeds and the like.

Duncan, Williams, Zehr, and Widholm, Planta (1985) 165:322-332 reflectsthat 97% of the plants cultured that produced callus were capable ofplant regeneration. Subsequent experiments with both inbreds and hybridsproduced 91% regenerable callus that produced plants. In a further studyin 1988, Songstad, Duncan & Widholm in Plant Cell Reports (1988),7:262-265 reports several media additions that enhance regenerability ofcallus of two inbred lines. Other published reports also indicated that“nontraditional” tissues are capable of producing somatic embryogenesisand plant regeneration. K. P. Rao, et al., Maize Genetics CooperationNewsletter, 60:64-65 (1986), refers to somatic embryogenesis from glumecallus cultures and B. V. Conger, et al., Plant Cell Reports, 6:345-347(1987) indicates somatic embryogenesis from the tissue cultures of maizeleaf segments. Thus, it is clear from the literature that the state ofthe art is such that these methods of obtaining plants are, and were,“conventional” in the sense that they are routinely used and have a veryhigh rate of success.

Tissue culture procedures of maize are described in Green and Rhodes,“Plant Regeneration in Tissue Culture of Maize,” Maize for BiologicalResearch (Plant Molecular Biology Association, Charlottesville, Va.1982, at 367-372) and in Duncan, et al., “The Production of CallusCapable of Plant Regeneration from Immature Embryos of Numerous Zea maysGenotypes,” 165 Planta 322-332 (1985). Thus, another aspect of thisinvention is to provide cells that upon growth and differentiationproduce maize plants having the physiological and morphologicalcharacteristics of inbred maize line 413A. In a preferred embodiment,cells of inbred maize line 413A are transformed genetically, for examplewith one or more genes described above, for example by using atransformation method described in U.S Pat. No. 6,114,608, andtransgenic plants of inbred maize line 413A are obtained and used forthe production of hybrid maize plants.

Maize is used as human food, livestock feed, and as raw material inindustry. Sweet corn kernels having a relative moisture of approximately72% are consumed by humans and may be processed by canning or freezing.The food uses of maize, in addition to human consumption of maizekernels, include both products of dry- and wet-milling industries. Theprincipal products of maize dry milling are grits, meal and flour. Themaize wet-milling industry can provide maize starch, maize syrups, anddextrose for food use. Maize oil is recovered from maize germ, which isa by-product of both dry- and wet-milling industries.

Maize, including both grain and non-grain portions of the plant, is alsoused extensively as livestock feed, primarily for beef cattle, dairycattle, hogs, and poultry. Industrial uses of maize include productionof ethanol, maize starch in the wet-milling industry and maize flour inthe dry-milling industry. The industrial applications of maize starchand flour are based on functional properties, such as viscosity, filmformation, adhesive properties, and ability to suspend particles. Themaize starch and flour have application in the paper and textileindustries. Other industrial uses include applications in adhesives,building materials, foundry binders, laundry starches, explosives,oil-well muds, and other mining applications. Plant parts other than thegrain of maize are also used in industry: for example, stalks and husksare made into paper and wallboard and cobs are used for fuel and to makecharcoal.

The seed of inbred maize line 413A or of inbred maize line 413A furthercomprising one or more single gene traits, the plant produced from theinbred seed, the hybrid maize plant produced from the crossing of theinbred, hybrid seed, and various parts of the hybrid maize plant can beutilized for human food, livestock feed, and as a raw material inindustry.

The present invention therefore also discloses an agricultural productcomprising a plant of the present invention or derived from a plant ofthe present invention. The present invention also discloses anindustrial product comprising a plant of the present invention orderived from a plant of the present invention. The present inventionfurther discloses methods of producing an agricultural or industrialproduct comprising planting seeds of the present invention, growingplant from such seeds, harvesting the plants and processing them toobtain an agricultural or industrial product.

DEPOSIT

Applicants have made a deposit of at least 2500 seeds of Inbred MaizeLine 413A with the American Type Culture Collection (ATCC), Manassas,Va., 20110-2209 U.S.A., ATCC Deposit No: PTA-4601. This deposit of theInbred Maize Line 413A will be maintained in the ATCC depository, whichis a public depository, for a period of 30 years, or 5 years after themost recent request, or for the effective life of the patent, whicheveris longer, and will be replaced if it becomes nonviable during thatperiod. Additionally, Applicants have satisfied all the requirements of37 C.F.R. §§1.801-1.809, including providing an indication of theviability of the sample. Applicants impose no restrictions on theavailability of the deposited material from the ATCC; however,Applicants have no authority to waive any restrictions imposed by law onthe transfer of biological material or its transportation in commerce.Applicants do not waive any infringement of its rights granted underthis patent or under the Plant Variety Protection Act (7 USC 2321 etseq.).

The foregoing invention has been described in detail by way ofillustration and example for purposes of clarity and understanding.However, it will be obvious that certain changes and modifications suchas single gene modifications and mutations, somaclonal variants, variantindividuals selected from large populations of the plants of the instantinbred and the like may be practiced within the scope of the invention,as limited only by the scope of the appended claims.

1-27. (canceled)
 28. An F1 hybrid maize seed produced by the methodcomprising crossing a first parent maize plant with a second parentmaize plant and harvesting the resultant first generation maize seed,wherein said first or second parent maize plant is a plant of inbredmaize inbred line 413A, seed of said line having been deposited underATCC Accession No: PTA-4601.
 29. An F1 hybrid plant grown from the seedof claim
 28. 30. An F1 hybrid maize seed according to claim 28, whereinsaid hybrid maize seed is a seed of hybrid GH5704, seed of said hybridhaving been deposited under ATCC Accession No: PTA-7221.
 31. An F1hybrid plant grown from the seed of claim 30.